Identification of Plasma Free Fatty Acid Carrier Proteins in Mice

Free fatty acid (FFA) transport through the bloodstream had been believed to mainly depend on binding to albumin. However, we recently discovered that albumin knockout (Alb^-/-) mice still have meaningful levels of FFAs in the bloodstream, indicating the presence of other FFA carrier proteins in plasma. To identify these proteins, we collected plasma from wildtype (WT) and Alb^-/- mice. Plasma proteins that can bind to FFAs were isolated by a fatty acid pulldown assay and identified by proteomic analysis. Additionally, plasma proteins were separated by fast protein liquid chromatography (FPLC), and the FPLC fractions were used to measure FFAs, cholesterol, triacylglycerol, and plasma proteins of interest. Furthermore, in silico docking simulations were used to identify potential FFA binding sites on the more highly expressed proteins isolated by the fatty acid pulldown assay to ascertain the likelihood that these proteins may have binding sites for FFA. Proteomic analysis on proteins isolated by the pulldown assay detected 202 proteins, collectively termed the FFA transportome. Examples of highly expressed proteins that showed FFA binding in the assay include albumin, transferrin, and apolipoprotein A-I. Notably, FFA levels in the FPLC fractions positively correlated with albumin, transferrin, HDL cholesterol, and Apo AI levels. The docking results further suggested that indeed many of the discovered proteins may have FFA binding sites. In conclusion, we elucidated the FFA transportome in mice, and this collection of FFA carrier proteins may facilitate FFA transport through the bloodstream along with albumin.